Enhancing autophagy by down-regulating GSK-3ß alleviates cisplatin-induced ototoxicity in vivo and in vitro.

Previous study reported that either selective GSK-3ß inhibitor or up-regulating autophagy can alleviate cisplatin-induced ototoxicity. Other studies indicate that the activity of GSK-3ß is closely associated with the autophagy level. The purpose of this study is to primarily explore the role of autophagy in the alleviation effect of GSK-3ß inhibition on cisplatin-induced ototoxicity in vivo and in vitro. We observed the autophagy changes induced by GSK-3ß inhibitor in outer hair cells (OHCs) in a cisplatin-induced ototoxicity rat model. In addition, autophagy inhibitor 3-MA was used in vitro experiments to observe the influence of autophagy inhibition on the cell protection effect due to GSK-3ß inactivation. The relationship among autophagy, GSK-3ß and cell damage were inferred. Negative regulation of GSK-3ß significantly enhanced autophagy and alleviated cisplatin-induced hearing loss, OHC death in vivo and apoptosis in vitro. The autophagy inhibitor 3-MA inverted the protective effect of negative regulation of GSK-3ß. These results indicated that enhancing autophagy may be a key downstream effect of GSK-3ß inhibition in the alleviation of cisplatin-induced ototoxicity both in vivo and in vitro.

Forskolin protects against cisplatin-induced ototoxicity by inhibiting apoptosis and ROS production.

Cisplatin is widely used in the treatment of various types of cancer. However, it could cause severe side effects such as ototoxicity, which greatly limit the clinical application of cisplatin. Forskolin (FSK) is a diterpene derived from the plant Coleus forskohlii, and has been proven an effective drug for cardiovascular disease, diabetes, and asthma because of its anti-oxidant and anti-inflammatory action. Here, we investigated the effects of FSK in cisplatin-induced ototoxicity, and we found that FSK could significantly protect against cisplatin-induced ototoxicity in both cell line and isolated mouse cochlear. Pretreatment of FSK attenuated cisplatin-induced hearing loss especially at high frequency regions. FSK inhibited the activation of mitochondrial apoptotic pathway as well as reactive oxygen species (ROS) production. Moreover, we identified PKA and MAPK signaling pathway which may be connected with the protective effect of FSK. Our study provided the first evidence that FSK may be used as a drug to weaken the ototoxicity induced by cisplatin.

Lactate dehydrogenase isoenzyme patterns in auricular pseudocyst fluid.

OBJECTIVE: We investigated lactate dehydrogenase isoenzyme patterns in the cyst fluid of auricular pseudocysts and autogenous blood, to assist the diagnosis of auricular pseudocyst. METHODS: Twenty patients with auricular pseudocysts participated in this study conducted in Kaohsiung Medical University Hospital between February 2007 and June 2010. Patterns of lactate dehydrogenase in cyst fluid and autogenous blood were analysed. RESULTS: Levels of lactate dehydrogenase 1 and 2 were lower in auricular pseudocysts than in autogenous blood, whereas levels of lactate dehydrogenase 4 and 5 were higher; this difference was statistically significant (p < 0.001). CONCLUSION: Lactate dehydrogenase isoenzyme patterns in auricular pseudocyst fluid indicated higher percentage distributions of lactate dehydrogenase 4 and 5 and lower percentage distributions of lactate dehydrogenase 1 and 2. An effective laboratory method of evaluating the different lactate dehydrogenase isoenzyme components was developed; this method may improve the accuracy of auricular pseudocyst diagnosis.

Matrix metalloproteinases and their inhibitors in non-neoplastic otorhinolaryngological disease.

Matrix metalloproteinases (MMPs) are a family of zinc and calcium-dependent endopeptidases that play a key role in extracellular matrix (ECM) degradation. MMPs are known to be important in normal remodelling processes. Overexpression and activation of MMPs or an imbalance of active MMPs and tissue inhibitors of metalloproteinases (TIMPs) has been linked with a number of specific disease states associated with the breakdown and remodelling of the extracellular matrix. MMPs and TIMPs play a role in the development and progression of conditions such as acute and chronic otitis media, nasal polyposis and Sjogren's disease of salivary glands. Their role in allergic rhinitis has not been proven although they do appear to have a role in asthma, a condition closely linked to rhinitis. The use of a broad spectrum MMP inhibitor has been shown to alter the outcome of acute otitis media and otitis media with effusion. Therapeutic strategies with anti-MMP molecules are currently being developed and may play a role in modulating the course of non-neoplastic otorhinolaryngological disease in the future.

Lactate dehydrogenase isozymes in and intralesional steroid injection therapy for pseudocyst of the auricle.

BACKGROUND: Various treatments for pseudocyst of the auricle have been reported. Recently, several patients have been treated successfully with steroid injections. The lactate dehydrogenase (LDH) levels of cystic fluid have been reported to be high, and the LDH-4 and LDH-5 isozymes were found to predominate, although the serum LDH levels and LDH isozyme patterns were normal. OBJECTIVE: To find a way of preventing auricular pseudocyst recurrences after steroid injection therapy and to examine the relationships between recurrences and the LDH level and isozyme pattern of the cystic fluid. METHODS: Steroid injection therapy was given to nine Japanese patients with pseudocyst of the auricle at our clinic between 1994 and 1999. We determined the LDH level and isozyme pattern of the cystic fluid and sera of seven patients. RESULTS: All nine patients in this series were treated successfully with steroid injections, although three suffered recurrences. Although the serum LDH levels of seven patients were almost normal, their cystic fluid LDH levels were very high. LDH-4 and LDH-5 predominated in the cystic fluid of all seven, although their serum LDH isozyme patterns were virtually normal. CONCLUSIONS: Auricular pseudocyst recurrences show no relationship with either the LDH levels or isozyme pattern. An undiluted solution of steroid fluid should be used in order to prevent recurrences.

Steroid injection therapy for pseudocyst of the auricle.

In our clinic, we encountered 8 patients with pseudocyst of the auricle during a 6-year period from 1987 to 1992 and injected steroid solution locally into the pseudocysts. Three of these patients had no recurrences and 4 were treated successfully after 1-3 recurrences. In only one case was this therapy ineffective. After 11 injections, which resulted in permanent deformity of the auricle, the patient underwent surgery. We believe that local steroid injection therapy should be the first choice method for treating auricular pseudocysts. However, frequent injections can cause auricular deformity and if there are more than 3 recurrences, the pseudocyst should be managed surgically. The lactate dehydrogenase levels of the cystic fluid were determined in 3 of the 8 patients and proved high; in only one patient was the etiology of the pseudocyst thought to be associated with minor trauma.

Pseudocyst of the auricle: case reports and its biochemical characteristics.

Pseudocyst of the auricle is a rare, asymptomatic cystic swelling of the auricle resulting from accumulation of yellow viscous fluid with unknown cause. We herein report 3 such patients in whom biochemical study of the aspirated fluid revealed markedly elevated activity of lactate dehydrogenase (LDH) and a preponderance of LDH-4 and LDH-5. We postulate that pseudocyst formation is due to the disruption of the auricular cartilage and that LDH in the fluid is released from the auricular cartilage. This observation supports that the lesions represent a pattern of chondromalacia. It is hypothesized that the cause of such pseudocysts is repeated minor trauma.

[Proteolytic and antiproteolytic activity of cholesteatoma and granulomatosis tissue of the middle ear]. / Aktywnosc proteolityczna i antyproteolityczna perlaka i ziarniny ucha srodkowego.

Cholesteatoma and granulomatosis tissue show proteolytic activity measured in pH 7.6. Cholesteatoma activity was found to be higher than this of granulomatosis tissue. Cholesteatoma does not appear to poses antiplasmin activity. Antiplasmin granulomatosis tissue activity is slight. Both cholesteatoma and granulomatosis tissue show antitrypsin activity. Antitrypsin cholesteatoma activity is lower than that of granulomatosis tissue.

[Cholesteatoma of the middle ear. An enzymatic study]. / Colesteatoma del oído medio. Estudio enzimático.

Forty specimens of aural cholesteatoma have been studied histoenzymological techniques. Acid phosphatases were the most representatives.

A comparative study of enzyme histochemical features in the gerbilline and human cholesteatoma.

Spontaneous and experimentally induced cholesteatoma in the Mongolian gerbil has been found to exhibit histopathological similarities to human aural cholesteatoma and has been suggested as an experimental model for studies of the clinical situation. In an attempt to further characterize this model, we compared experimentally induced cholesteatomas in the external auditory canal from gerbils with those of the human ear by means of a correlated histopathologic and enzyme histochemical study. The human and gerbilline cholesteatomas revealed similar histopathologic features. Even enzyme histochemically, the human and experimentally induced cholesteatomas demonstrated similar features. Thus glucose-6-phosphate dehydrogenase activity, an indicator of oxidative metabolism, was demonstrated especially in the stratum granulosum cells of the heavily orthokeratinizing squamous epithelium adjacent to the cholesteatomas. The human ear canal skin also revealed enzyme histochemical characteristics similar to the squamous epithelium lining the human cholesteatoma. The hydrolytic enzyme activity (leucyl-aminopeptidase) was strong in the connective tissue surrounding human cholesteatoma when compared with that of ear canal skin. In the gerbilline cholesteatoma, this activity was demonstrated especially in the connective tissue adjacent to eroded bone, which possibly may facilitate cholesteatoma progression. We conclude that experimentally induced cholesteatoma has both histophatological and enzyme histochemical similarities to human aural cholesteatoma and therefore it is suggested that the gerbilline model may be used for studies on the development of human cholesteatoma. Our results support the view that cholesteatoma may originate from migrated hyperkeratinizing cells from the epidermis of the tympanic membrane or the meatus.

Cathepsin activity in cholesteatoma.

Collagenolytic cathepsin, presumed to play an important role in bone destruction of cholesteatoma, was investigated in cholesteatoma epithelium, subepithelial granulation tissue, skin from the bony external auditory meatus and, temporal bone. The enzyme extracted from tissues was proven to be lysosomal cathepsin B by SDS gel electrophoresis in the use of human type I and type III collagen. alpha-N-benzoyl-DL-arginine-2-naphthylamide HCl (BANA) was supposed to be specific for cathepsin B, and so BANA-hydrolase activity was measured as collagen-degrading cathepsin. The results showed that tissues had cathepsin B with its optimal pH 6.0, and that cathepsin B activity revealed a significant increase in the subepithelial granulation tissue. A strong activity of acid phosphatase found in the subepithelial granulation tissue seems to reflect the existence of an active metabolism of substances in the granulation tissue. These findings suggest that collagen is resorbed in the subepithelial granulation tissue in the presence of cholesteatoma.

Collagenase activity in cholesteatoma.

Collagenase activity of cholesteatoma epithelium, temporal bone and ear canal skin was investigated. The enzyme extracted from these tissues was proved to be collagenase by disc electrophoresis. Collagenase activity was measured directly in homogenates of tissues. Activity was greatest in the skin. The activity in cholesteatomas was either higher than that of the bone, or the activity was higher in bone than in the cholesteatoma. These findings suggest that some variation of collagenase activity may be due to the stage of cholesteatoma, active or inactive in collagen metabolism of bone destruction, and that the ear canal skin plays a great role in bone destruction, this supporting the immigration theory.

Low plasma renin concentration in patients with Meniere's disease (preliminary report).

Endolymphatic hydrops is the histopathologic hallmark of Meniere's disease. The cause of excess inner ear fluid is unknown, but the possibility of a systemic abnormality of fluid-solute regulation has been considered. One indicator of systemic fluid and electrolyte balance is the enzyme renin. Plasma renin levels were measured in patients with a clinical diagnosis of Meniere's disease and compared to a group of patients who did not have Meniere's disease. The mean plasma renin concentration of patients with Meniere's disease was statistically lower than that of the other subjects. The study's results are consistent with a systemic abnormality in fluid and solute regulation in some patients with M eniere's disease, leading to abnormal expansion of the extracellular fluid volume, and possibly contributing to endolympathic hydrops.

Localization of collagenase in human middle ear cholesteatoma.

The process of connective tissue breakdown in chronic otitis media is described in the context of recent advances in our understanding of collagen degradation and bone resorption. The significance of the initial step in collagen breakdown, brought about by the action of a specific collagen dissolving enzyme is emphasized in terms of recent studies in other chronic inflammatory diseases characterized by connective tissue breakdown. Bone resorption, a characteristic feature of chronic otitis media, requires the breakdown of collagen, which comprises over 90 percent of bone protein. Evidence in support of collagenase in bone resorption from adjacent tissue (in this case, inflammatory connective tissue) would require identification of the enzyme in cells involved in the inflammatory process adjacent to the resorbing bone. Collagenase was found localized in frozen sections of canal wall skin, middle ear granulation and in cholesteatoma by a specific binding of the enzyme with an antiserum produced against purified human skin collagenase. The antigen antibody complex was labelled with fluorescein. Collagenase appeared in the subepithelial connective tissue of cholesteatoma, granulation tissue from the middle ear and the dermis of canal skin; but was not seen in the keratin layer, epithelium or the epidermal appendages. The enzyme appeared within certain fibroblasts, macrophages and endothelial cells of capillary buds. Collagenase enhanced by chronic inflammation attacks the intact collagen molecule, making it susceptible to further digestion by other proteases that are also products of inflammation. This process brings about resorption of connective tissue and bone.